P1287: DISTINGUISHING BETWEEN PRIMARY MEDIASTINAL LARGE B-CELL LYMPHOMA AND DIFFUSE LARGE B-CELL LYMPHOMA USING THE LYMPH3CX GENE EXPRESSION PROFILING ASSAY
Authors: R. S. Robetorye, C. A. Ramsower, C. L. McKinney, L. M. Rimsza
Journal: HemaSphere (2022)

Background: Primary mediastinal large B-cell lymphoma (PMBL) is a distinct lymphoid neoplasm arising in the mediastinum that is morphologically similar to diffuse large B-cell lymphoma (DLBCL), but has distinct clinical, immunophenotypic, genotypic, and molecular features. Although differentiation of PMBL from DLBCL can be important for guiding subsequent therapy, interpretation of morphologic findings and associated immunohistochemical stains in small, and often crushed, mediastinal biopsies can be difficult. Aims: Therefore, we developed and fully validated a clinical gene expression profiling assay in the Molecular Diagnostics Arizona Laboratory (MDAZL) using the Lymph3Cx gene expression signature to distinguish these lymphomas using formalin-fixed paraffin-embedded (FFPE) tissue specimens. This assay includes and builds upon the Lymph2Cx expression signature and can subtype DLBCL into germinal center B-cell (GCB)-type, activated B-cell (ABC)-type, and unclassifiable (UNC) categories as well as accurately and robustly differentiate PMBL from DLBCL using the NanoString nCounter® platform. Methods: A hematopathologist identifies a large B-cell lymphoma, and additional testing is ordered for further classification of the lymphoma (including Lymph3Cx analysis). Pathological assessment of tumor content is performed on the H&E-stained slide, and unstained sections are sent to MDAZL for further processing. Total RNA is isolated and quantitated in MDAZL, and the RNA is hybridized overnight to fluorescently labeled probes in the Lymph3Cx assay panel. The probe/RNA complexes are quantitated on a NanoString nCounter Digital Analyzer. Algorithmic analysis is performed (R-script) for quality control (QC) and subtype determination and resulting data are verified by the pathologist and loaded directly into the EMR for correlation with clinical data and final diagnosis. Results: So far, 40 clinical specimens have been subjected to Lymph3Cx testing in the MDAZL Laboratory with the following results: PMBL: 19 (47.5%); DLBCL: 15 (37.5%); Unclear: 5 (12.5%); Failed QC: 1 (2.5%). Of the 15 cases with gene expression signatures consistent with DLBCL, cell-of-origin results showed the following results: GCB: 8 (53.33%); ABC: 5 (33.33%); UNC: 2 (13.33%). Ordering locations included Mayo Clinic Arizona: 14 (35%); Mayo Clinic Minnesota: 7 (17.5%); Mayo Clinic Florida: 5 (12.5%); other external sites: 14 (35%). Average in-laboratory turn-around-time for these cases was 1.8 days, with an average of 3.3 days from sample receipt to final report. Summary/Conclusion: The Molecular Diagnostics–Arizona Laboratory (MDAZL) has performed a full clinical validation of the performance characteristics of the Lymph3Cx assay and is the first molecular diagnostics laboratory to offer the Lymph3Cx assay as a clinical molecular diagnostic test for distinguishing PMBL from DLBCL and molecular cell-of-origin subtyping of DLBCL to patients both inside and outside of the Mayo Clinic. With a relatively rapid turn-around-time, the Lymph3Cx assay is suitable for use in routine hematopathology practice for distinguishing PMBL and DLBCL and is also suitable for definitive identification of PMBL cases for possible use in ongoing clinical trials to identify more specific therapeutics for treatment of PMBL.